FoxM1抑制剂硫链丝菌素对髓母细胞瘤Daoy细胞顺铂化疗敏感性的影响

Effect of FoxM1 inhibitor thiostrepton on CDDP chemosensitivity in medulloblastoma Daoy cells

目的研究硫链丝菌素通过抑制FoxMl表达对髓母细胞瘤Daoy细胞顺铂（cDDP）化疗敏感性的影响。方法体外常规培养髓母细胞瘤Daoy细胞株，CCK8法检测1、1．25、1．5、1．75、2、3、5μmol／L硫链丝菌素，1、2、3、4、5、10、20、40μmol／LcDDP及1μmol／L硫链丝菌素与1、2、3、4、5、10、20、40μmol／LcDDP联合处理后对细胞的抑制率．金式公式分析两药间的互相作用：根据分析结果采用1μmol／L硫链丝菌素、3μmol／LcDDP单独或联合作用Daoy细胞24h，流式细胞仪检测细胞凋亡的变化，Westernblotting检测FoxMl及凋亡相关蛋白bcl-2、bax、caspase3、聚腺苷二磷酸核糖聚合酶（PARP）表达的变化。结果硫链丝菌素可浓度依赖性地抑制髓母细胞瘤Daoy细胞的生长。金式公式分析显示硫链丝菌素与低浓度（1、2、3、4、5μmol／L）cDDP存在明显的协同作用；流式细胞分析仪显示硫链丝菌素单药组、cDDP单药组细胞凋亡率高于对照组，而联合用药组细胞凋产率高于硫链丝菌素单药组和cDDP单药组，差异有统计学意义（P〈0．05）。Westemblowing结果显示，与对照组比较，硫链丝菌素单药组和联合用药组FoxMl蛋白表达明显下降，差异有统计学意义（P〈0．05）；与硫链丝菌素单药组、cDDP单药组比较，联合用药组细胞bcl-2蛋白表达明显下降，而bax、caspase3、PARP蛋白表达明显升高。结论硫链丝菌素和cDDP在低浓度下具有明显的协同抗髓母细胞瘤作用，其机制可能与硫链丝菌素抑制FoxMl表达后增强了cDDP诱导凋广的能力有关。

Objective To investigate the effects of FoxM1 inhibition by thiostrepton on CDDP chemosensitivity in medulloblastoma Daoy cells. Methods Medulloblastoma Daoy cells were cultured in vitro; the inhibitory rates of Daoy cells after the treatments of 1, 1.25, 1.5, 1.75, 2, 3, 5 p.mol/L thiostrepton （thiostrepton treatment groups）, or 1, 2, 3, 4, 5, 10, 20 and 40 μmol/L CDDP （CDDP treatment groups） or 1 μmol/L thiostrepton combined with 1, 2, 3, 4, 5, 10, 20 and 40 μmol/L cDDP （combination treatment groups） were evaluated by CCK8 assay, and the interaction between thiostrepton and cDDP was analyzed by means of Jin＇s formulatin. According to the analysis results, Daoy cells were treated with thiostrepton （1 μmol/L） and CDDP （3 p, mol/L） combination or either drug alone for 24 h, and then, flow cytometry was used to observe the changes of apoptosis; Western blotting was employed to detect the expressions of FoxM1 and apoptosis-relating proteins of bcl-2, bax, caspase 3 and poly-ADP-ribose polymerase （PARP）. Results Thiostrepton and cDDP used alone could inhibit the proliferation of Daoy cells, and there was simple addition or synergistic effect after combined treatment （1, 2, 3, 4 and 5 μmol/L cDDP and thiostrepton）. Flow cytometric analysis showed that the apoptosis rate of thiostrepton treatment groups and CDDP treatment groups was significantly higher than that of control groups, while that of combination treatment groups was significantly higher than that of thiostreptontreatment groups and CDDP treatment groups （P〈0.05）. Western blotting indicated that the FoxM1 expression level in the combination treatment groups and thiostrepton treatment groups was significantly lower than that in the control group （P〈0.05）; as compared with that in the thiostrepton treatment groups and CDDP treatment groups, the bcl-2 protein level was significantly lower in the combination treatment groups, while Bax, caspase 3 and PARP protein expressions were significantly increased （P〈0.05）. Conclusion Thiostrepton acts synergistically with cDDP in low concentration, and its mechanism may involve in inhibiting FoxM1 expression, and then, enhancing cDDP-induced apoptosis.